Labeled antisense oligonucleotides will be prepared and used as essential tools in studies of the kinetics of cellular uptake and relationship between oligonucleotide modification and efficacy of its transport through cellular membranes. Effective chemical method for an attachment of 14C, 3H or 35S radiolabel to phosphorothioate monoesters or diesters via alkylation procedure using 14CH3I and C3H3I or other radioactive reporting group, or via oxidation of phosphate triesters or amidoesters with 35S will be developed. Oligothymidylic acids length of 4 and 20 nucleoside units, labeled in terminal phosphorothioate group with 14C, 3H or 35S will be synthesized. They will bear one or more modifications such as methylphosphonates, hydroxymethylphosphonates and carboranylmethylphosphonates. Steroselectivity of cellular uptake of all-Rp and all-Sp methylphosphonate oligonucleotides will be studied using radiolabeled oligonucleotide models. Following radiolabeled carboranyl pyrimidine nucleosides and oligonucleotides designed for use in BNCT and as antiviral agents will be synthesized: [6-14C] 5- carboranyl-2'-deoxyuridine, [6-14C] 5-carboranyl-2',3'-dideoxy-3- thiacytidine, [3H and/or 14C] 5'-CDU-dd(T)4, [3H and/or 14C] 5'-d (T)4 CDU, [3H and/or 14C] 5'-D (t)2-CDU-d(T)2, [3H and/or 14C] 5'-CDU-d(T)3- CDU, [3H and/or 14C] 5'- (T)5 . Synthesis from labeled monomers, or post-synthesis labeling via carboranyl-lithium intermediates will be used. General method for direct tritiation of unmodified oligonucleotides based on carefully controlled heterogeneous metal catalyzed exchange with tritium gas will be developed. PROPOSED COMMERCIAL APPLICATION: Most of the compounds developed in this project will be offered to the research community in Moravek Biochemicals commercial catalogs. New general technology will be utilized in custom synthesis services of Moravek Biochemicals and in the expansion of the oligonucleotide product line.